RNA-Protein Biochemistry – As an RNA-protein lab, we do a lot of ‘basic’ biochemistry everyday. Ranging from gel shifts to gel filtration to fluorescence polarization assays, we routinely use these techniques to ask questions like the binding kinetics and specificity of the RNA-binding proteins that we love to study.

Fluorescence Spectroscopy – To study ribosome assembly in vitro with single-intermediate resolution, we have built a 2-photon microscope with single-molecule sensitivity. Adaptations of Fluctuation Correlation Spectroscopy & Coincidence Analysis allow us to extract kinetic traces with ~1 second resolution over the duration of the hour-long assembly reaction. We have also designed and built an automated microfluidic reactor which can precisely prepare and initiate the requisite large number of reactions. We are presently investigating late stages of 30S assembly in the 3’ major domain using this novel technique.

Isothermal Titration Calorimetry – We use ITC to measure the binding constants of protein to protein (e.g. the formation of a tight heterodimer between the ribosomal protein S6 and S18, as shown above) and protein to RNA complexes.

ITC Mixing Capillary

X-Ray Crystallography – Nothing says so much about a protein-RNA complex as an elegant crystal structure. We use crystallography to help us visualize the interactions between proteins and RNA of large complexes (e.g. the 30S ribosome sub-domain).

NMR Spectroscopy – While it does not produce pretty crystals, arguably NMR is the best tool to study flexible molecules in solution, and we routinely employ NMR to examine (and even solve!) the structures of our RNA and proteins.

Mass Spectroscopy – More recently, we have delved into mass spectroscopy and developed methods to monitor the different rates of isotope labeling of ribosomal proteins from in vitro reconstitution experiments. Using a novel Fourier-based peak fitting algorithm, we are able to precisely measure the binding rates of all 20 ribosomal proteins of the 30S subunit.